Age-related hearing loss (ARHL), the most prevalent form of hearing loss in humans, is commonly caused by degenerative changes (or biological aging) and apoptosis of the cochlear inner and outer hair cells (IHCs, OHCs). Despite the fact that ARHL has been extensively studied using various mouse models, the causes of hair cell aging and apoptosis are still largely unknown. Our long-term goal is to examine the cellular and molecular mechanisms of biological aging of IHCs and OHCs. In the first part of the proposal we will attempt to answer two critical questions of how cochlear hair cells undergo aging, and why hair cell degeneration proceeds from the basal to the apical turn and from OHCs to IHCs. In the second part, we will focus on the molecular mechanisms of why OHCs are more susceptible to aging or other insults than IHCs by determining the roles of Clu, a gene that is differentially expressed in OHCs. A battery of electrophysiological, immunohistochemical, advanced imaging, and molecular biological techniques will be used to examine cellular changes in ultrastructure and mechanical and electrophysiological properties of IHCs and OHCs in the base and apex of the cochlea, respectively, in CBA/J strain mice during aging. Our hair cell-specific RNA-seq experiments using IHCs and OHCs obtained from younger and aging mice will enable us to reveal molecular mechanisms by identifying genes underlying age-related changes in IHCs and OHCs. We have three specific aims. Aim 1 is to examine the changes in mechanotransduction and mechanical and electrophysiological properties of hair cells during aging. Aim 2 is to examine the changes in gene expression underlying molecular mechanisms of aging of IHCs and OHCs. Aim 3 is to test the hypothesis that the elevated levels of Clu expression in OHCs relative to IHCs play a key role in the OHC?s selective susceptibility to aging. Completion of these aims is critical to understand the intrinsic cellular and molecular mechanisms of hair cell aging. The knowledge gained from this research is also essential for identifying targets for therapeutic intervention that aims to decelerate ARHL.